Identification of potential prognosticators for sepsis through expression analysis of transcriptomic data from sepsis survivors and nonsurvivors

Background@#Infection can be severely complicated by a dysregulated, whole-body inflammatory response known as sepsis. While previous research showed that genetic predisposition is linked to outcome differences, current patient characterization fails to determine which septic patients have greater tendencies to develop into severe sepsis or go into septic shock. As such, the identification of prognostic biomarkers may assist in identifying these high-risk patients and help improve the clinical management of the disease.@*Objective@#In this study, we aimed to identify molecular patterns involved in sepsis. We also aimed to identify essential genes associated with the disease’s survival which could serve as potential prognosticators for the disease. @*Methods@#We used weighted gene co-expression analysis (WGCNA) to analyze GSE63042, an RNA expression dataset from 129 patients with systemic inflammatory response syndrome or sepsis, including 78 sepsis survivors and 28 sepsis nonsurvivors. This analysis included identifying gene modules that differentiate sepsis survivors from nonsurvivors and qualitatively assessing differentially expressed genes. We then used STRING’s protein-protein interaction and gene ontology analysis to determine the functional and pathway relationships of the genes in the top modules. Lastly, we assessed the prognosticator abilities of the hub genes using ROC analysis. @*Results@#We found four diverse co-expression gene modules significantly associated with sepsis survival. Our differential gene expression analysis, combined with protein-protein interaction and gene ontology analysis, revealed that the hub genes of these modules – TAF10, SNAPIN, PSME2, PSMB9, JUNB, and CEBPD – may serve as candidate markers for sepsis prognosis. These markers were significantly downregulated in sepsis nonsurvivors compared with sepsis survivors.@*Conclusion@#Weighted gene co-expression analysis, gene ontology enrichment analysis, and proteinprotein network interaction analysis of transcriptomic data from sepsis survivors and nonsurvivors revealed TAF10, SNAPIN, PSME2, PSMB9, JUNB, and CEBPD as potential biomarkers for sepsis prognosis. These genes are associated with functions related to proper immune response, and their downregulation in sepsis nonsurvivors suggests eventual immune exhaustion in late sepsis. Further analyses, however, are necessary to validate their roles in sepsis progression and patient survival.

Host protein Snapin interacts with human cytomegalovirus pUL130 and affects viral DNA replication.

The interplay between the host and Human cytomegalovirus (HCMV) plays a pivotal role in the outcome of an infection. HCMV growth in endothelial and epithelial cells requires expression of viral proteins UL128, UL130, and UL131 proteins (UL128-131), of which UL130 is the largest gene and the only one that is not interrupted by introns.Mutation of the C terminus of the UL130 protein causes reduced tropism of endothelial cells (EC). However, very few host factors have been identified that interact with the UL130 protein. In this study, HCMV UL130 protein was shown to directly interact with the human protein Snapin in human embryonic kidney HEK293 cells by Yeast two-hybrid screening, in vitro glutathione S-transferase (GST) pull-down, and co-immunoprecipitation. Additionally, heterologous expression of protein UL130 revealed co-localization with Snapin in the cell membrane and cytoplasm of HEK293 cells using fluorescence confocal microscopy. Furthermore, decreasing the level of Snapin via specific small interfering RNAs decreased the number of viral DNA copies and titer inHCMV-infected U373-S cells. Taken together, these results suggest that Snapin, the pUL130 interacting protein, has a role in modulating HCMV DNA synthesis.

LRRK2 phosphorylates Snapin and inhibits interaction of Snapin with SNAP-25

Leucine-rich repeat kinase 2 (LRRK2) is a gene that, upon mutation, causes autosomal-dominant familial Parkinson's disease (PD). Yeast two-hybrid screening revealed that Snapin, a SNAP-25 (synaptosomal-associated protein-25) interacting protein, interacts with LRRK2. An in vitro kinase assay exhibited that Snapin is phosphorylated by LRRK2. A glutathione-S-transferase (GST) pull-down assay showed that LRRK2 may interact with Snapin via its Ras-of-complex (ROC) and N-terminal domains, with no significant difference on interaction of Snapin with LRRK2 wild type (WT) or its pathogenic mutants. Further analysis by mutation study revealed that Threonine 117 of Snapin is one of the sites phosphorylated by LRRK2. Furthermore, a Snapin T117D phosphomimetic mutant decreased its interaction with SNAP-25 in the GST pull-down assay. SNAP-25 is a component of the SNARE (Soluble NSF Attachment protein REceptor) complex and is critical for the exocytosis of synaptic vesicles. Incubation of rat brain lysate with recombinant Snapin T117D, but not WT, protein caused decreased interaction of synaptotagmin with the SNARE complex based on a co-immunoprecipitation assay. We further found that LRRK2-dependent phosphorylation of Snapin in the hippocampal neurons resulted in a decrease in the number of readily releasable vesicles and the extent of exocytotic release. Combined, these data suggest that LRRK2 may regulate neurotransmitter release via control of Snapin function by inhibitory phosphorylation.